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Updated: Aug 8, 2026
An Efficient and High Yield Method for Isolation of Mouse Dendritic Cell Subsets
Published on: April 18, 2016
Dendritic cells that process and present nominal antigens to naive T lymphocytes are derived from CD2+ precursors
M Takamizawa1, A Rivas, F Fagnoni
1Department of Pediatrics, Tokyo University Hospital, Japan.
Insights
Researchers discovered two distinct types of dendritic cells (DCs) derived from CD2+ and CD2- precursors. Only CD2+ derived DCs can present soluble antigens to naive T cells, highlighting distinct precursor roles in immune responses.
Area of Science:
- Immunology
- Cell Biology
Background:
- Dendritic cells (DCs) are potent antigen-presenting cells (APCs) crucial for initiating adaptive immune responses.
- Mature DCs exhibit distinct morphology, lack lineage markers, and express high levels of MHC and costimulatory molecules.
- Previous methods for DC preparation yielded cells with varying functional capabilities.
Purpose of the Study:
- To investigate the functional differences between dendritic cells (DCs) derived from peripheral blood mononuclear cells (PBMCs) with and without prior CD2+ cell depletion.
- To identify distinct precursor populations within PBMCs responsible for generating functionally different DCs.
- To characterize the phenotypic and functional properties of CD2+ and CD2- DC precursors.
Main Methods:
- Comparison of DC preparation methods involving CD2+ cell depletion from PBMCs.
- Morphological analysis (light and electron microscopy) of isolated DC precursor populations.
- Functional assays including mixed lymphocyte reaction (MLR) and T cell sensitization to soluble antigens.
- Flow cytometry to assess the expression of cell surface markers (e.g., CD54, CD86) on DC precursors.
- Inhibition studies using antibodies targeting T cell and DC surface molecules.
Main Results:
- DCs derived from CD2+ depleted PBMCs were functionally distinct from those derived from un-depleted PBMCs.
- Two precursor populations, CD2+ and CD2-, were identified within HLA-DRbright, lineage-negative PBMCs.
- Both precursor populations generated DCs that stimulated T cell proliferation in MLR and autologous T cell responses to recall antigens after culture.
- Only DCs derived from CD2+ precursors could sensitize naive T cells to soluble antigens (e.g., keyhole limpet hemocyanin, HIV gp160).
- Expression of CD54 and CD86 increased significantly on CD2+ DC precursors after culture and differentiation, unlike CD2- precursors.
Conclusions:
- Peripheral blood contains at least two functionally distinct DC precursor populations: CD2+ and CD2-.
- CD2+ DC precursors are essential for generating APCs capable of presenting soluble antigens to naive CD4+ T cells.
- These findings reveal distinct roles for phenotypically different precursors in generating functionally specialized dendritic cells.
Abstract:
Dendritic cells (DC) are potent APC that, in mature form, can be distinguished from other mononuclear cells on the basis of their distinct morphology, absence of lineage markers, and dense expression of MHC and costimulatory molecules. While comparing different DC preparation methods, we observed that DC derived from cultured PBMC that had been depleted of CD2+ cells before culture were functionally distinct from DC derived from PBMC that had not been depleted of CD2+ cells. Thus, both types of DC stimulated allogeneic T cells to proliferate in the MLR, but only DC derived from CD2+ precursors could sensitize naive T cells to soluble Ags such as keyhole limpet hemocyanin and HIV gp160 glycoprotein. Subsequent studies confirmed the existence of CD2+ and CD2- DC precursor populations among HLA-DRbright, lineage-negative PBMC. Immediately after their isolation, these populations were morphologically similar to one another by light and electron microscopy, and neither had substantial Ag-presenting activity. After culture for 24 to 48 h with supernatant from PHA-activated PBMC, both populations developed dendrites, formed clusters with T cells, and stimulated allogeneic T cell responses in the MLR as well as autologous T cell responses to tetanus toxoid, a recall Ag. However, CD2+ DC precursors alone gave rise to APC that presented soluble Ags to naive CD4+ T cells, a property that could be inhibited by Abs to CD4, CD11a, and CD28 on T cells or CD86 on DC. The expression of CD54 and CD86 on CD2+ DC precursors was increased markedly after their culture and differentiation, while the expression of these molecules on CD2- DC precursors was not remarkably changed. These findings reveal the existence of two functionally distinct populations of DC, each derived from a phenotypically distinct precursor present in monocyte-depleted peripheral blood.
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